Proteome Birthdating: A Single-Sample Approach for Measuring Global Turnover Dynamics and “Protein Age”
Within a cell, proteins have distinct and highly variable half-lives. As a result, the molecular ages of proteins can range from seconds to years. How the age of a protein influences its environmental interactions is a largely unexplored area of biology. To facilitate such studies, we recently devel...
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| Format: | Article |
| Language: | English |
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Bio-protocol LLC
2025-05-01
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| Series: | Bio-Protocol |
| Online Access: | https://bio-protocol.org/en/bpdetail?id=5296&type=0 |
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| author | Michael Meadow Sarah Broas Margaret Hoare Maria Ahmed Fatemeh Alimohammadi Kevin Welle Kyle Swovick Jennifer Hryhorenko Anushka Jain John Martinez Andrei Seluanov Vera Gorbunova Abigail Buchwalter Sina Ghaemmaghami |
| author_facet | Michael Meadow Sarah Broas Margaret Hoare Maria Ahmed Fatemeh Alimohammadi Kevin Welle Kyle Swovick Jennifer Hryhorenko Anushka Jain John Martinez Andrei Seluanov Vera Gorbunova Abigail Buchwalter Sina Ghaemmaghami |
| author_sort | Michael Meadow |
| collection | DOAJ |
| description | Within a cell, proteins have distinct and highly variable half-lives. As a result, the molecular ages of proteins can range from seconds to years. How the age of a protein influences its environmental interactions is a largely unexplored area of biology. To facilitate such studies, we recently developed a technique termed “proteome birthdating” that differentially labels proteins based on their time of synthesis. Proteome birthdating enables analyses of age distributions of the proteome by tandem mass spectrometry (LC–MS/MS) and provides a methodology for investigating the protein age selectivity of diverse cellular pathways. Proteome birthdating can also provide measurements of protein turnover kinetics from single, sequentially labeled samples. Here, we provide a practical guide for conducting proteome birthdating in in vitro model systems. The outlined workflow covers cell culture, isotopic labeling, protein extraction, enzymatic digestion, peptide cleanup, mass spectrometry, data processing, and theoretical considerations for interpretation of the resulting data. |
| format | Article |
| id | doaj-art-8d2cd747d7224629844f042d7b83a3bf |
| institution | Kabale University |
| issn | 2331-8325 |
| language | English |
| publishDate | 2025-05-01 |
| publisher | Bio-protocol LLC |
| record_format | Article |
| series | Bio-Protocol |
| spelling | doaj-art-8d2cd747d7224629844f042d7b83a3bf2025-08-20T03:48:46ZengBio-protocol LLCBio-Protocol2331-83252025-05-0115910.21769/BioProtoc.5296Proteome Birthdating: A Single-Sample Approach for Measuring Global Turnover Dynamics and “Protein Age”Michael Meadow0Sarah Broas1Margaret Hoare2Maria Ahmed3Fatemeh Alimohammadi4Kevin Welle5Kyle Swovick6Jennifer Hryhorenko7Anushka Jain8John Martinez9Andrei Seluanov10Vera Gorbunova11Abigail Buchwalter12Sina Ghaemmaghami13Department of Biology, University of Rochester, NY, USAMedical Scientist Training Program, University of Rochester, NY, USADepartment of Biology, University of Rochester, NY, USADepartment of Biology, University of Rochester, NY, USADepartment of Biology, University of Rochester, NY, USADepartment of Pharmacology and Physiology, University of Rochester Medical Center, Rochester, NY, USAUniversity of Rochester Mass Spectrometry Resource Laboratory, NY, USAUniversity of Rochester Mass Spectrometry Resource Laboratory, NY, USAUniversity of Rochester Mass Spectrometry Resource Laboratory, NY, USADepartment of Biology, University of Rochester, NY, USADepartment of Biology, University of Rochester, NY, USADepartment of Biology, University of Rochester, NY, USADepartment of Medicine, University of Rochester Medical Center, Rochester, NY, USADepartment of Biology, University of Rochester, NY, USADepartment of Medicine, University of Rochester Medical Center, Rochester, NY, USACardiovascular Research Institute, University of California San Francisco, San Francisco, CA, USADepartment of Biology, University of Rochester, NY, USAUniversity of Rochester Mass Spectrometry Resource Laboratory, NY, USAWithin a cell, proteins have distinct and highly variable half-lives. As a result, the molecular ages of proteins can range from seconds to years. How the age of a protein influences its environmental interactions is a largely unexplored area of biology. To facilitate such studies, we recently developed a technique termed “proteome birthdating” that differentially labels proteins based on their time of synthesis. Proteome birthdating enables analyses of age distributions of the proteome by tandem mass spectrometry (LC–MS/MS) and provides a methodology for investigating the protein age selectivity of diverse cellular pathways. Proteome birthdating can also provide measurements of protein turnover kinetics from single, sequentially labeled samples. Here, we provide a practical guide for conducting proteome birthdating in in vitro model systems. The outlined workflow covers cell culture, isotopic labeling, protein extraction, enzymatic digestion, peptide cleanup, mass spectrometry, data processing, and theoretical considerations for interpretation of the resulting data.https://bio-protocol.org/en/bpdetail?id=5296&type=0 |
| spellingShingle | Michael Meadow Sarah Broas Margaret Hoare Maria Ahmed Fatemeh Alimohammadi Kevin Welle Kyle Swovick Jennifer Hryhorenko Anushka Jain John Martinez Andrei Seluanov Vera Gorbunova Abigail Buchwalter Sina Ghaemmaghami Proteome Birthdating: A Single-Sample Approach for Measuring Global Turnover Dynamics and “Protein Age” Bio-Protocol |
| title | Proteome Birthdating: A Single-Sample Approach for Measuring Global Turnover Dynamics and “Protein Age” |
| title_full | Proteome Birthdating: A Single-Sample Approach for Measuring Global Turnover Dynamics and “Protein Age” |
| title_fullStr | Proteome Birthdating: A Single-Sample Approach for Measuring Global Turnover Dynamics and “Protein Age” |
| title_full_unstemmed | Proteome Birthdating: A Single-Sample Approach for Measuring Global Turnover Dynamics and “Protein Age” |
| title_short | Proteome Birthdating: A Single-Sample Approach for Measuring Global Turnover Dynamics and “Protein Age” |
| title_sort | proteome birthdating a single sample approach for measuring global turnover dynamics and protein age |
| url | https://bio-protocol.org/en/bpdetail?id=5296&type=0 |
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